Biotechnology potentially offers unconventional routes to new cancer therapies, and research in this area has focused on using viruses as vectors to deliver therapeutic genes to tumours. Unfortunately, serious issues with the safety of viral vectors have been encountered (1, 2) and only recently has a virus system showing good specificity and tumour infiltration been described (3). Several types of bacteria have also been investigated as delivery vectors, and as therapeutics in their own right (4). Spores of some species of Clostridium, which are strictly anaerobic bacteria, naturally target tumours with high specificity following intravenous administration, because the dormant spores can germinate and grow only in the hypoxic cores of solid tumours (5-7) which are difficult to target using viral vectors (8, 9). The growing bacteria secrete proteases inside the tumour, rapidly digesting the tumour mass. This approach is especially interesting because it directly targets the hypoxic cells in poorly vascular regions which tend to be refractory to conventional treatments. Any Clostridium cells entering normal tissue from a colonized tumour would be poisoned by oxygen and die. Stringent spacial containment by oxygen is an excellent safety feature absent from treatments using aerobic bacteria (10, 11).
Augmentation of the anti-tumour effect of Clostridium strains has been sought by genetically modifying them to express therapeutic proteins, mainly enzymes which sensitize the tumour to certain chemotherapeutic agents (13-17). The ‘prodrug’ CB1954 (5-(aziridin-1-yl)-2,4-dinitrobenzamide) shown in FIG. 1 is an attractive candidate because its clinical safety has been demonstrated (18) and a 10,000-fold increase in toxicity is achieved upon its activation by a suitable nitroreductase (NTR) enzyme to the 4-hydroxylamine (4HX) derivative (5-(aziridin-1-yl)-4-hydroxylamino-2-nitrobenzamide) which is a DNA cross-linking agent (19).